Structural basis of GSDME pore formation and its regulation by S-palmitoylation

• 成果类型: Article
• 署名作者: Du, Gang; Ehrmann, Julian F.; Lieberman, Judy; Wu, Hao
• 署名单位: Harvard University; Harvard Medical School; Harvard University; Harvard University Medical Affiliates; Boston Children's Hospital; Program in Cellular & Molecular Medicine (PCMM); Harvard University; Harvard Medical School
• 刊物名称: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• ISSN/ISSBN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.2614339123
• 发表日期: 2026-09-01
• 页码: e2614339123
• 关键词: pyroptosis; gasdermin E; cryo-em structure; s-palmitoylation; cancer-associated mutations; gasdermin d; inflammatory caspases; pyroptosis; cleavage

摘要

Pyroptosis is defined as gasdermin-mediated lytic programmed cell death. Gasdermin E (GSDME), a substrate of the apoptotic caspase-3, can convert apoptosis into pyroptosis, with critical roles in antitumor immunity and chemotherapy-induced tissue damage. Despite its importance, the structural mechanism of GSDME pore formation and its regulation by posttranslational modifications remain largely unknown. Here, we present the cryo-electron microscopy (cryo-EM) structure at 3.16 & Aring; resolution of the human GSDME N-terminal (NT) pore using proteins expressed from mammalian cells. The structure reveals a GSDME-NT pore assembled mainly as a 28-subunit homo-oligomer, and a dramatic conformational rearrangement from the autoinhibited state, with refolding of the two beta-hairpins in each monomer to form a membrane-spanning beta-barrel with an acidic conduit. Unexpectedly, we identify endogenous S-palmitoylation of C45, C168, and C180, required for membrane binding and pore formation. In addition, extra cryo-EM densities are visible adjacent to the C45 side chain, potentially corresponding to the flexibly linked palmitate chain. Structure-guided mutagenesis demonstrates that these palmitoylation sites synergistically control pore formation. The structure served as a molecular blueprint for analyzing cancer-associated mutations, known to disrupt GSDME function. These mutations cluster at functional hotspots in the oligomerization interfaces, membrane-contact regions, and the beta-barrel, where they disrupt pore integrity. Collectively, our findings establish palmitoylation as an obligatory licensing step for membrane binding and pore formation, provide structural visualization of a palmitoylated gasdermin, and reveal how cancer-associated mutations impair pyroptotic function. This structure and these insights will be useful for developing strategies that target GSDME to treat cancer and inflammatory disease.